The long-term objective of this research work is to gain a better understanding of the mechanism of protein synthesis initiation in red blood cells. This study will include the following: (1) Peptide Chain Initiation Factors - We have isolated four protein factors, EIF-1, Co-EIF-1, EIF-2 and EIF-3 and have demonstrated their requirement in protein synthesis initiation in reticulocytes. We propose to further purify these factors using conventional methods and study their precise roles in the specific steps in the initiation process. (2) Protein Synthesis Inhibitors - We will study the precise roles of three protein factors: HCR, Fx and Met-tRNAf deacylase in the regulation of protein synthesis in red blood cells. These three factors have been reported to inhibit protein synthesis in vitro. We will also study the mechanism of protein synthesis inhibition by antibiotics such as pactamycin. In our study, we will use several specific and sensitive peptide chain initiation assay methods recently developed in our laboratory.